Biallelic CRELD1 variants cause a multisystem syndrome, including neurodevelopmental phenotypes, cardiac dysrhythmias, and frequent infections.

PURPOSE: We sought to delineate a multisystem disorder caused by recessive cysteine-rich with epidermal growth factor-like domains 1 (CRELD1) gene variants. METHODS: The impact of CRELD1 variants was characterized through an international collaboration utilizing next-generation DNA sequencing, gene knockdown, and protein overexpression in Xenopus tropicalis, and in vitro analysis of patient immune cells. RESULTS: Biallelic variants in CRELD1 were found in 18 participants from 14 families. Affected individuals displayed an array of phenotypes involving developmental delay, early-onset epilepsy, and hypotonia, with about half demonstrating cardiac arrhythmias and some experiencing recurrent infections. Most harbored a frameshift in trans with a missense allele, with 1 recurrent variant, p.(Cys192Tyr), identified in 10 families. X tropicalis tadpoles with creld1 knockdown displayed developmental defects along with increased susceptibility to induced seizures compared with controls. Additionally, human CRELD1 harboring missense variants from affected individuals had reduced protein function, indicated by a diminished ability to induce craniofacial defects when overexpressed in X tropicalis. Finally, baseline analyses of peripheral blood mononuclear cells showed similar proportions of immune cell subtypes in patients compared with healthy donors. CONCLUSION: This patient cohort, combined with experimental data, provide evidence of a multisystem clinical syndrome mediated by recessive variants in CRELD1.

SRSF1 haploinsufficiency is responsible for a syndromic developmental disorder associated with intellectual disability.

SRSF1 (also known as ASF/SF2) is a non-small nuclear ribonucleoprotein (non-snRNP) that belongs to the arginine/serine (R/S) domain family. It recognizes and binds to mRNA, regulating both constitutive and alternative splicing. The complete loss of this proto-oncogene in mice is embryonically lethal. Through international data sharing, we identified 17 individuals (10 females and 7 males) with a neurodevelopmental disorder (NDD) with heterozygous germline SRSF1 variants, mostly de novo, including three frameshift variants, three nonsense variants, seven missense variants, and two microdeletions within region 17q22 encompassing SRSF1. Only in one family, the de novo origin could not be established. All individuals featured a recurrent phenotype including developmental delay and intellectual disability (DD/ID), hypotonia, neurobehavioral problems, with variable skeletal (66.7%) and cardiac (46%) anomalies. To investigate the functional consequences of SRSF1 variants, we performed in silico structural modeling, developed an in vivo splicing assay in Drosophila, and carried out episignature analysis in blood-derived DNA from affected individuals. We found that all loss-of-function and 5 out of 7 missense variants were pathogenic, leading to a loss of SRSF1 splicing activity in Drosophila, correlating with a detectable and specific DNA methylation episignature. In addition, our orthogonal in silico, in vivo, and epigenetics analyses enabled the separation of clearly pathogenic missense variants from those with uncertain significance. Overall, these results indicated that haploinsufficiency of SRSF1 is responsible for a syndromic NDD with ID due to a partial loss of SRSF1-mediated splicing activity.

Bi-allelic TTI1 variants cause an autosomal-recessive neurodevelopmental disorder with microcephaly.

Telomere maintenance 2 (TELO2), Tel2 interacting protein 2 (TTI2), and Tel2 interacting protein 1 (TTI1) are the three components of the conserved Triple T (TTT) complex that modulates activity of phosphatidylinositol 3-kinase-related protein kinases (PIKKs), including mTOR, ATM, and ATR, by regulating the assembly of mTOR complex 1 (mTORC1). The TTT complex is essential for the expression, maturation, and stability of ATM and ATR in response to DNA damage. TELO2- and TTI2-related bi-allelic autosomal-recessive (AR) encephalopathies have been described in individuals with moderate to severe intellectual disability (ID), short stature, postnatal microcephaly, and a movement disorder (in the case of variants within TELO2). We present clinical, genomic, and functional data from 11 individuals in 9 unrelated families with bi-allelic variants in TTI1. All present with ID, and most with microcephaly, short stature, and a movement disorder. Functional studies performed in HEK293T cell lines and fibroblasts and lymphoblastoid cells derived from 4 unrelated individuals showed impairment of the TTT complex and of mTOR pathway activity which is improved by treatment with Rapamycin. Our data delineate a TTI1-related neurodevelopmental disorder and expand the group of disorders related to the TTT complex.

Intranasal Carbetocin Reduces Hyperphagia, Anxiousness, and Distress in Prader-Willi Syndrome: CARE-PWS Phase 3 Trial.

CONTEXT: Prader-Willi syndrome (PWS) is a rare genetic disorder characterized by endocrine and neuropsychiatric problems including hyperphagia, anxiousness, and distress. Intranasal carbetocin, an oxytocin analog, was investigated as a selective oxytocin replacement therapy. OBJECTIVE: To evaluate safety and efficacy of intranasal carbetocin in PWS. DESIGN: Randomized, double-blind, placebo-controlled phase 3 trial with long-term follow-up. SETTING: Twenty-four ambulatory clinics at academic medical centers. PARTICIPANTS: A total of 130 participants with PWS aged 7 to 18 years. INTERVENTIONS: Participants were randomized to 9.6 mg/dose carbetocin, 3.2 mg/dose carbetocin, or placebo 3 times daily during an 8-week placebo-controlled period (PCP). During a subsequent 56-week long-term follow-up period, placebo participants were randomly assigned to 9.6 mg or 3.2 mg carbetocin, with carbetocin participants continuing at their previous dose. MAIN OUTCOME MEASURES: Primary endpoints assessed change in hyperphagia (Hyperphagia Questionnaire for Clinical Trials [HQ-CT]) and obsessive-compulsive symptoms (Children's Yale-Brown Obsessive-Compulsive Scale [CY-BOCS]) during the PCP for 9.6 mg vs placebo, and the first secondary endpoints assessed these same outcomes for 3.2 mg vs placebo. Additional secondary endpoints included assessments of anxiousness and distress behaviors (PWS Anxiousness and Distress Behaviors Questionnaire [PADQ]) and clinical global impression of change (CGI-C). RESULTS: Because of onset of the COVID-19 pandemic, enrollment was stopped prematurely. The primary endpoints showed numeric improvements in both HQ-CT and CY-BOCS which were not statistically significant; however, the 3.2-mg arm showed nominally significant improvements in HQ-CT, PADQ, and CGI-C scores vs placebo. Improvements were sustained in the long-term follow-up period. The most common adverse event during the PCP was mild to moderate flushing. CONCLUSIONS: Carbetocin was well tolerated, and the 3.2-mg dose was associated with clinically meaningful improvements in hyperphagia and anxiousness and distress behaviors in participants with PWS. CLINICAL TRIALS REGISTRATION NUMBER: NCT03649477.

Recurring germline mosaicism in a family due to reversion of an inherited derivative chromosome 8 from an 8;21 translocation with interstitial telomeric sequences.

BACKGROUND: Mosaicism for chromosomal structural abnormalities, other than marker or ring chromosomes, is rarely inherited. METHODS: We performed cytogenetics studies and breakpoint analyses on a family with transmission of mosaicism for a derivative chromosome 8 (der(8)), resulting from an unbalanced translocation between the long arms of chromosomes 8 and 21 over three generations. RESULTS: The proband and his maternal half-sister had mosaicism for a der(8) cell line leading to trisomy of the distal 21q, and both had Down syndrome phenotypic features. Mosaicism for a cell line with the der(8) and a normal cell line was also detected in a maternal half-cousin. The der(8) was inherited from the maternal grandmother who had four abnormal cell lines containing the der(8), in addition to a normal cell line. One maternal half-aunt had the der(8) and an isodicentric chromosome 21 (idic(21)). Sequencing studies revealed microhomologies at the junctures of the der(8) and idic(21) in the half-aunt, suggesting a replicative mechanism in the rearrangement formation. Furthermore, interstitial telomeric sequences (ITS) were identified in the juncture between chromosomes 8 and 21 in the der(8). CONCLUSION: Mosaicism in the proband, his half-sister and half-cousin resulting from loss of chromosome 21 material from the der(8) appears to be a postzygotic event due to the genomic instability of ITS and associated with selective growth advantage of normal cells. The reversion of the inherited der(8) to a normal chromosome 8 in this family resembles revertant mosaicism of point mutations. We propose that ITS could mediate recurring revertant mosaicism for some constitutional chromosomal structural abnormalities.

Novel variants in KAT6B spectrum of disorders expand our knowledge of clinical manifestations and molecular mechanisms.

The phenotypic variability associated with pathogenic variants in Lysine Acetyltransferase 6B (KAT6B, a.k.a. MORF, MYST4) results in several interrelated syndromes including Say-Barber-Biesecker-Young-Simpson Syndrome and Genitopatellar Syndrome. Here we present 20 new cases representing 10 novel KAT6B variants. These patients exhibit a range of clinical phenotypes including intellectual disability, mobility and language difficulties, craniofacial dysmorphology, and skeletal anomalies. Given the range of features previously described for KAT6B-related syndromes, we have identified additional phenotypes including concern for keratoconus, sensitivity to light or noise, recurring infections, and fractures in greater numbers than previously reported. We surveyed clinicians to qualitatively assess the ways families engage with genetic counselors upon diagnosis. We found that 56% (10/18) of individuals receive diagnoses before the age of 2 years (median age = 1.96 years), making it challenging to address future complications with limited accessible information and vast phenotypic severity. We used CRISPR to introduce truncating variants into the KAT6B gene in model cell lines and performed chromatin accessibility and transcriptome sequencing to identify key dysregulated pathways. This study expands the clinical spectrum and addresses the challenges to management and genetic counseling for patients with KAT6B-related disorders.

Pathogenic MAST3 Variants in the STK Domain Are Associated with Epilepsy.

OBJECTIVE: The MAST family of microtubule-associated serine-threonine kinases (STKs) have distinct expression patterns in the developing and mature human and mouse brain. To date, only MAST1 has been conclusively associated with neurological disease, with de novo variants in individuals with a neurodevelopmental disorder, including a mega corpus callosum. METHODS: Using exome sequencing, we identify MAST3 missense variants in individuals with epilepsy. We also assess the effect of these variants on the ability of MAST3 to phosphorylate the target gene product ARPP-16 in HEK293T cells. RESULTS: We identify de novo missense variants in the STK domain in 11 individuals, including 2 recurrent variants p.G510S (n = 5) and p.G515S (n = 3). All 11 individuals had developmental and epileptic encephalopathy, with 8 having normal development prior to seizure onset at <2 years of age. All patients developed multiple seizure types, 9 of 11 patients had seizures triggered by fever and 9 of 11 patients had drug-resistant seizures. In vitro analysis of HEK293T cells transfected with MAST3 cDNA carrying a subset of these patient-specific missense variants demonstrated variable but generally lower expression, with concomitant increased phosphorylation of the MAST3 target, ARPP-16, compared to wild-type. These findings suggest the patient-specific variants may confer MAST3 gain-of-function. Moreover, single-nuclei RNA sequencing and immunohistochemistry shows that MAST3 expression is restricted to excitatory neurons in the cortex late in prenatal development and postnatally. INTERPRETATION: In summary, we describe MAST3 as a novel epilepsy-associated gene with a potential gain-of-function pathogenic mechanism that may be primarily restricted to excitatory neurons in the cortex. ANN NEUROL 2021;90:274-284.

Metabolic impact of pathogenic variants in the mitochondrial glutamyl-tRNA synthetase EARS2.

Glutamyl-tRNA synthetase 2 (encoded by EARS2) is a mitochondrial aminoacyl-tRNA synthetase required to translate the 13 subunits of the electron transport chain encoded by the mitochondrial DNA. Pathogenic EARS2 variants cause combined oxidative phosphorylation deficiency, subtype 12 (COXPD12), an autosomal recessive disorder involving lactic acidosis, intellectual disability, and other features of mitochondrial compromise. Patients with EARS2 deficiency present with variable phenotypes ranging from neonatal lethality to a mitigated disease with clinical improvement in early childhood. Here, we report a neonate homozygous for a rare pathogenic variant in EARS2 (c.949G>T; p.G317C). Metabolomics in primary fibroblasts from this patient revealed expected abnormalities in TCA cycle metabolites, as well as numerous changes in purine, pyrimidine, and fatty acid metabolism. To examine genotype-phenotype correlations in COXPD12, we compared the metabolic impact of reconstituting these fibroblasts with wild-type EARS2 versus four additional EARS2 variants from COXPD12 patients with varying clinical severity. Metabolomics identified a group of signature metabolites, mostly from the TCA cycle and amino acid metabolism, that discriminate between EARS2 variants causing relatively mild and severe COXPD12. Taken together, these findings indicate that metabolomics in patient-derived fibroblasts may help establish genotype-phenotype correlations in EARS2 deficiency and likely other mitochondrial disorders.

Variants in the SK2 channel gene (KCNN2) lead to dominant neurodevelopmental movement disorders.

KCNN2 encodes the small conductance calcium-activated potassium channel 2 (SK2). Rodent models with spontaneous Kcnn2 mutations show abnormal gait and locomotor activity, tremor and memory deficits, but human disorders related to KCNN2 variants are largely unknown. Using exome sequencing, we identified a de novo KCNN2 frameshift deletion in a patient with learning disabilities, cerebellar ataxia and white matter abnormalities on brain MRI. This discovery prompted us to collect data from nine additional patients with de novo KCNN2 variants (one nonsense, one splice site, six missense variants and one in-frame deletion) and one family with a missense variant inherited from the affected mother. We investigated the functional impact of six selected variants on SK2 channel function using the patch-clamp technique. All variants tested but one, which was reclassified to uncertain significance, led to a loss-of-function of SK2 channels. Patients with KCNN2 variants had motor and language developmental delay, intellectual disability often associated with early-onset movement disorders comprising cerebellar ataxia and/or extrapyramidal symptoms. Altogether, our findings provide evidence that heterozygous variants, likely causing a haploinsufficiency of the KCNN2 gene, lead to novel autosomal dominant neurodevelopmental movement disorders mirroring phenotypes previously described in rodents.

De novo heterozygous missense and loss-of-function variants in CDC42BPB are associated with a neurodevelopmental phenotype.

CDC42BPB encodes MRCKß (myotonic dystrophy-related Cdc42-binding kinase beta), a serine/threonine protein kinase, and a downstream effector of CDC42, which has recently been associated with Takenouchi-Kosaki syndrome, an autosomal dominant neurodevelopmental disorder. We identified 12 heterozygous predicted deleterious variants in CDC42BPB (9 missense, 2 frameshift, and 1 nonsense) in 14 unrelated individuals (confirmed de novo in 11/14) with neurodevelopmental disorders including developmental delay/intellectual disability, autism, hypotonia, and structural brain abnormalities including cerebellar vermis hypoplasia and agenesis/hypoplasia of the corpus callosum. The frameshift and nonsense variants in CDC42BPB are expected to be gene-disrupting and lead to haploinsufficiency via nonsense-mediated decay. All missense variants are located in highly conserved and functionally important protein domains/regions: 3 are found in the protein kinase domain, 2 are in the citron homology domain, and 4 in a 20-amino acid sequence between 2 coiled-coil regions, 2 of which are recurrent. Future studies will help to delineate the natural history and to elucidate the underlying biological mechanisms of the missense variants leading to the neurodevelopmental and behavioral phenotypes.

The CHD4-related syndrome: a comprehensive investigation of the clinical spectrum, genotype-phenotype correlations, and molecular basis.

PURPOSE: Sifrim-Hitz-Weiss syndrome (SIHIWES) is a recently described multisystemic neurodevelopmental disorder caused by de novo variants inCHD4. In this study, we investigated the clinical spectrum of the disorder, genotype-phenotype correlations, and the effect of different missense variants on CHD4 function. METHODS: We collected clinical and molecular data from 32 individuals with mostly de novo variants in CHD4, identified through next-generation sequencing. We performed adenosine triphosphate (ATP) hydrolysis and nucleosome remodeling assays on variants from five different CHD4 domains. RESULTS: The majority of participants had global developmental delay, mild to moderate intellectual disability, brain anomalies, congenital heart defects, and dysmorphic features. Macrocephaly was a frequent but not universal finding. Additional common abnormalities included hypogonadism in males, skeletal and limb anomalies, hearing impairment, and ophthalmic abnormalities. The majority of variants were nontruncating and affected the SNF2-like region of the protein. We did not identify genotype-phenotype correlations based on the type or location of variants. Alterations in ATP hydrolysis and chromatin remodeling activities were observed in variants from different domains. CONCLUSION: The CHD4-related syndrome is a multisystemic neurodevelopmental disorder. Missense substitutions in different protein domains alter CHD4 function in a variant-specific manner, but result in a similar phenotype in humans.

Correction: The CHD4-related syndrome: a comprehensive investigation of the clinical spectrum, genotype-phenotype correlations, and molecular basis.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Recurrent arginine substitutions in the ACTG2 gene are the primary driver of disease burden and severity in visceral myopathy.

Visceral myopathy with abnormal intestinal and bladder peristalsis includes a clinical spectrum with megacystis-microcolon intestinal hypoperistalsis syndrome and chronic intestinal pseudo-obstruction. The vast majority of cases are caused by dominant variants in ACTG2; however, the overall genetic architecture of visceral myopathy has not been well-characterized. We ascertained 53 families, with visceral myopathy based on megacystis, functional bladder/gastrointestinal obstruction, or microcolon. A combination of targeted ACTG2 sequencing and exome sequencing was used. We report a molecular diagnostic rate of 64% (34/53), of which 97% (33/34) is attributed to ACTG2. Strikingly, missense mutations in five conserved arginine residues involving CpG dinucleotides accounted for 49% (26/53) of disease in the cohort. As a group, the ACTG2-negative cases had a more favorable clinical outcome and more restricted disease. Within the ACTG2-positive group, poor outcomes (characterized by total parenteral nutrition dependence, death, or transplantation) were invariably due to one of the arginine missense alleles. Analysis of specific residues suggests a severity spectrum of p.Arg178>p.Arg257>p.Arg40 along with other less-frequently reported sites p.Arg63 and p.Arg211. These results provide genotype-phenotype correlation for ACTG2-related disease and demonstrate the importance of arginine missense changes in visceral myopathy.

Mutation update for the SATB2 gene.

SATB2-associated syndrome (SAS) is an autosomal dominant neurodevelopmental disorder caused by alterations in the SATB2 gene. Here we present a review of published pathogenic variants in the SATB2 gene to date and report 38 novel alterations found in 57 additional previously unreported individuals. Overall, we present a compilation of 120 unique variants identified in 155 unrelated families ranging from single nucleotide coding variants to genomic rearrangements distributed throughout the entire coding region of SATB2. Single nucleotide variants predicted to result in the occurrence of a premature stop codon were the most commonly seen (51/120 = 42.5%) followed by missense variants (31/120 = 25.8%). We review the rather limited functional characterization of pathogenic variants and discuss current understanding of the consequences of the different molecular alterations. We present an expansive phenotypic review along with novel genotype-phenotype correlations. Lastly, we discuss current knowledge of animal models and present future prospects. This review should help provide better guidance for the care of individuals diagnosed with SAS.

De novo and inherited TCF20 pathogenic variants are associated with intellectual disability, dysmorphic features, hypotonia, and neurological impairments with similarities to Smith-Magenis syndrome.

BACKGROUND: Neurodevelopmental disorders are genetically and phenotypically heterogeneous encompassing developmental delay (DD), intellectual disability (ID), autism spectrum disorders (ASDs), structural brain abnormalities, and neurological manifestations with variants in a large number of genes (hundreds) associated. To date, a few de novo mutations potentially disrupting TCF20 function in patients with ID, ASD, and hypotonia have been reported. TCF20 encodes a transcriptional co-regulator structurally related to RAI1, the dosage-sensitive gene responsible for Smith-Magenis syndrome (deletion/haploinsufficiency) and Potocki-Lupski syndrome (duplication/triplosensitivity). METHODS: Genome-wide analyses by exome sequencing (ES) and chromosomal microarray analysis (CMA) identified individuals with heterozygous, likely damaging, loss-of-function alleles in TCF20. We implemented further molecular and clinical analyses to determine the inheritance of the pathogenic variant alleles and studied the spectrum of phenotypes. RESULTS: We report 25 unique inactivating single nucleotide variants/indels (1 missense, 1 canonical splice-site variant, 18 frameshift, and 5 nonsense) and 4 deletions of TCF20. The pathogenic variants were detected in 32 patients and 4 affected parents from 31 unrelated families. Among cases with available parental samples, the variants were de novo in 20 instances and inherited from 4 symptomatic parents in 5, including in one set of monozygotic twins. Two pathogenic loss-of-function variants were recurrent in unrelated families. Patients presented with a phenotype characterized by developmental delay, intellectual disability, hypotonia, variable dysmorphic features, movement disorders, and sleep disturbances. CONCLUSIONS: TCF20 pathogenic variants are associated with a novel syndrome manifesting clinical characteristics similar to those observed in Smith-Magenis syndrome. Together with previously described cases, the clinical entity of TCF20-associated neurodevelopmental disorders (TAND) emerges from a genotype-driven perspective.

Correction to: De novo and inherited TCF20 pathogenic variants are associated with intellectual disability, dysmorphic features, hypotonia, and neurological impairments with similarities to Smith-Magenis syndrome.

It was highlighted that the original article [1] contained a typographical error in the Results section. Subject 17 was incorrectly cited as Subject 1. This Correction article shows the revised statement. The original article has been updated.

Cornelia de Lange syndrome in diverse populations.

Cornelia de Lange syndrome (CdLS) is a dominant multisystemic malformation syndrome due to mutations in five genes-NIPBL, SMC1A, HDAC8, SMC3, and RAD21. The characteristic facial dysmorphisms include microcephaly, arched eyebrows, synophrys, short nose with depressed bridge and anteverted nares, long philtrum, thin lips, micrognathia, and hypertrichosis. Most affected individuals have intellectual disability, growth deficiency, and upper limb anomalies. This study looked at individuals from diverse populations with both clinical and molecularly confirmed diagnoses of CdLS by facial analysis technology. Clinical data and images from 246 individuals with CdLS were obtained from 15 countries. This cohort included 49% female patients and ages ranged from infancy to 37 years. Individuals were grouped into ancestry categories of African descent, Asian, Latin American, Middle Eastern, and Caucasian. Across these populations, 14 features showed a statistically significant difference. The most common facial features found in all ancestry groups included synophrys, short nose with anteverted nares, and a long philtrum with thin vermillion of the upper lip. Using facial analysis technology we compared 246 individuals with CdLS to 246 gender/age matched controls and found that sensitivity was equal or greater than 95% for all groups. Specificity was equal or greater than 91%. In conclusion, we present consistent clinical findings from global populations with CdLS while demonstrating how facial analysis technology can be a tool to support accurate diagnoses in the clinical setting. This work, along with prior studies in this arena, will assist in earlier detection, recognition, and treatment of CdLS worldwide.

LINE- and Alu-containing genomic instability hotspot at 16q24.1 associated with recurrent and nonrecurrent CNV deletions causative for ACDMPV.

Transposable elements modify human genome by inserting into new loci or by mediating homology-, microhomology-, or homeology-driven DNA recombination or repair, resulting in genomic structural variation. Alveolar capillary dysplasia with misalignment of pulmonary veins (ACDMPV) is a rare lethal neonatal developmental lung disorder caused by point mutations or copy-number variant (CNV) deletions of FOXF1 or its distant tissue-specific enhancer. Eighty-five percent of 45 ACDMPV-causative CNV deletions, of which junctions have been sequenced, had at least one of their two breakpoints located in a retrotransposon, with more than half of them being Alu elements. We describe a novel ∼35 kb-large genomic instability hotspot at 16q24.1, involving two evolutionarily young LINE-1 (L1) elements, L1PA2 and L1PA3, flanking AluY, two AluSx, AluSx1, and AluJr elements. The occurrence of L1s at this location coincided with the branching out of the Homo-Pan-Gorilla clade, and was preceded by the insertion of AluSx, AluSx1, and AluJr. Our data show that, in addition to mediating recurrent CNVs, L1 and Alu retrotransposons can predispose the human genome to formation of variably sized CNVs, both of clinical and evolutionary relevance. Nonetheless, epigenetic or other genomic features of this locus might also contribute to its increased instability.

Autosomal recessive Noonan syndrome associated with biallelic LZTR1 variants.

PURPOSE: To characterize the molecular genetics of autosomal recessive Noonan syndrome. METHODS: Families underwent phenotyping for features of Noonan syndrome in children and their parents. Two multiplex families underwent linkage analysis. Exome, genome, or multigene panel sequencing was used to identify variants. The molecular consequences of observed splice variants were evaluated by reverse-transcription polymerase chain reaction. RESULTS: Twelve families with a total of 23 affected children with features of Noonan syndrome were evaluated. The phenotypic range included mildly affected patients, but it was lethal in some, with cardiac disease and leukemia. All of the parents were unaffected. Linkage analysis using a recessive model supported a candidate region in chromosome 22q11, which includes LZTR1, previously shown to harbor mutations in patients with Noonan syndrome inherited in a dominant pattern. Sequencing analyses of 21 live-born patients and a stillbirth identified biallelic pathogenic variants in LZTR1, including putative loss-of-function, missense, and canonical and noncanonical splicing variants in the affected children, with heterozygous, clinically unaffected parents and heterozygous or normal genotypes in unaffected siblings. CONCLUSION: These clinical and genetic data confirm the existence of a form of Noonan syndrome that is inherited in an autosomal recessive pattern and identify biallelic mutations in LZTR1.

Identification of novel candidate disease genes from de novo exonic copy number variants.

BACKGROUND: Exon-targeted microarrays can detect small (<1000 bp) intragenic copy number variants (CNVs), including those that affect only a single exon. This genome-wide high-sensitivity approach increases the molecular diagnosis for conditions with known disease-associated genes, enables better genotype-phenotype correlations, and facilitates variant allele detection allowing novel disease gene discovery. METHODS: We retrospectively analyzed data from 63,127 patients referred for clinical chromosomal microarray analysis (CMA) at Baylor Genetics laboratories, including 46,755 individuals tested using exon-targeted arrays, from 2007 to 2017. Small CNVs harboring a single gene or two to five non-disease-associated genes were identified; the genes involved were evaluated for a potential disease association. RESULTS: In this clinical population, among rare CNVs involving any single gene reported in 7200 patients (11%), we identified 145 de novo autosomal CNVs (117 losses and 28 intragenic gains), 257 X-linked deletion CNVs in males, and 1049 inherited autosomal CNVs (878 losses and 171 intragenic gains); 111 known disease genes were potentially disrupted by de novo autosomal or X-linked (in males) single-gene CNVs. Ninety-one genes, either recently proposed as candidate disease genes or not yet associated with diseases, were disrupted by 147 single-gene CNVs, including 37 de novo deletions and ten de novo intragenic duplications on autosomes and 100 X-linked CNVs in males. Clinical features in individuals with de novo or X-linked CNVs encompassing at most five genes (224 bp to 1.6 Mb in size) were compared to those in individuals with larger-sized deletions (up to 5 Mb in size) in the internal CMA database or loss-of-function single nucleotide variants (SNVs) detected by clinical or research whole-exome sequencing (WES). This enabled the identification of recently published genes (BPTF, NONO, PSMD12, TANGO2, and TRIP12), novel candidate disease genes (ARGLU1 and STK3), and further confirmation of disease association for two recently proposed disease genes (MEIS2 and PTCHD1). Notably, exon-targeted CMA detected several pathogenic single-exon CNVs missed by clinical WES analyses. CONCLUSIONS: Together, these data document the efficacy of exon-targeted CMA for detection of genic and exonic CNVs, complementing and extending WES in clinical diagnostics, and the potential for discovery of novel disease genes by genome-wide assay.